Folate is required as 5-methyltetrahydrofolate (5-methylTHF) for methylation reactions such as DNA methylation, and as 5,10-methylenetetrahydrofolate (5,10-methyleneTHF) for conversion of deoxyuridylate into thymidylate, a required reaction for DNA synthesis. Various experiments indicate that deficiency of folate in either form, leading to abnormalities in DNA synthesis or in DNA methylation, could enhance colorectal carcinogenesis. Methylenetetrahydrofolate reductase (MTHFR) plays a central role in folate metabolism by irreversibly converting 5,10-methyleneTHF into 5-methylTHF. A single nucleotide polymorphism of the MTHFR gene (677C ->T; alanine-to-valine), correlates with reduced enzyme activity. Thus, MTHFR 677TT homozygotes tend to accumulate 5,10-methyleneTHF intracellularly at the expense of 5-methylTHF. MTHFR TT homozygotes appear to be hyper-responders to folate or alcohol, who are at relatively low risk (compared to those with CC or CT genotypes) if they have a "low-risk" diet (high folate, low alcohol), but may have elevated risks if they have a "high-risk" diet (high alcohol, low folate). Building upon data from 2 ongoing randomized clinical trials of folic acid and colorectal adenoma incidence, we propose to examine if blood and rectal measures of genomic DNA methylation and uracil incorporation influence risk of adenoma. We will then study if the MTHFR 677 genotype influences how nutritional factors (folate, alcohol, as well as vitamins B2, B6 and B 12) affects DNA methylation and uracil incorporation. Specifically, we hypothesize that MTHFR TT homozygotes are at elevated risk through genomic DNA hypomethylation (relative to the other genotypes) if they consume alcohol and low folate diets, but have lower uracil misincorporation by having more folate in the form required for DNA synthesis (5,10-methylene-THF). In addition to clarifying the mechanism by which folate influences risk of colorectal neoplasia, we anticipate these studies will resolve whether genomic hypomethylation and uracil misincorporation can be used as biomarkers of folate adequacy and adenoma risk.